Dosimetry for a study of low-dose radiation cataracts among Chernobyl clean-up workers.

A cohort of 8,607 Ukrainian Chernobyl clean-up workers during 1986-1987 was formed to study cataract formation after ionizing radiation exposure. Study eligibility required the availability of sufficient exposure information to permit the reconstruction of doses to the lens of the eye. Eligible groups included civilian workers, such as those who built the "sarcophagus" over the reactor, Chernobyl Nuclear Power Plant Workers, and military reservists who were conscripted for clean-up work. Many of the official doses for workers were estimates, because only a minority wore radiation badges. For 106 military workers, electron paramagnetic resonance (EPR) measurements of extracted teeth were compared with the recorded doses as the basis to adjust the recorded gamma-ray doses and provide estimates of uncertainties. Beta-particle doses to the lens were estimated with an algorithm devised to take into account the nature and location of Chernobyl work, time since the accident, and protective measures taken. A Monte Carlo routine generated 500 random estimates for each individual from the uncertainty distributions of the gamma-ray dose and of the ratio of beta-particle to gamma-ray doses. The geometric mean of the 500 combined beta-particle and gamma-ray dose estimates for each individual was used in the data analyses. The median estimated lens dose for the cohort was 123 mGy, while 4.4% received >500 mGy.

Cataracts among Chernobyl clean-up workers: implications regarding permissible eye exposures.

The eyes of a prospective cohort of 8,607 Chernobyl clean-up workers (liquidators) were assessed for cataract at 12 and 14 years after exposure. The prevalence of strictly age-related cataracts was low, as expected (only 3.9% had nuclear cataracts at either examination), since 90% of the cohort was younger than 55 years of age at first examination. However, posterior subcapsular or cortical cataracts characteristic of radiation exposure were present in 25% of the subjects. The data for Stage 1 cataracts, and specifically for posterior subcapsular cataracts, revealed a significant dose response. When various cataract end points were analyzed for dose thresholds, the confidence intervals all excluded values greater than 700 mGy. Linear-quadratic dose-response models yielded mostly linear associations, with weak evidence of upward curvature. The findings do not support the ICRP 60 risk guideline assumption of a 5-Gy threshold for "detectable opacities" from protracted exposures but rather point to a dose-effect threshold of under 1 Gy. Thus, given that cataract is the dose-limiting ocular pathology in current eye risk guidelines, revision of the allowable exposure of the human visual system to ionizing radiation should be considered.

Mice heterozygous for the ATM gene are more sensitive to both X-ray and heavy ion exposure than are wildtypes.

Previous studies have shown that the eyes of ATM heterozygous mice exposed to low-LET radiation (X-rays) are significantly more susceptible to the development of cataracts than are those of wildtype mice. The findings, as well as others, run counter to the assumption underpinning current radiation safety guidelines, that individuals are all equally sensitive to the biological effects of radiation. A question, highly relevant to human space activities is whether or not, in similar fashion there may exist a genetic predisposition to high-LET radiation damage. Mice haplodeficient for the ATM gene and wildtypes were exposed to 325 mGy of 1 GeV/amu 56Fe ions at the AGS facility of Brookhaven National Laboratory. The fluence was equivalent to 1 ion per lens epithelial cell nuclear area. Controls consisted of irradiated wildtype as well as unirradiated wildtype and heterozygous mice. Prevalence analyses for stage 0.5-3.0 cataracts indicated that not only cataract onset but also progression were accelerated in the mice haplo-deficient for the ATM gene. The data show that heterozygosity for the ATM gene predisposes the eye to the cataractogenic influence of heavy ions and suggest that ATM heterozygotes in the human population may also be radiosensitive. This may have to be considered in the selection of individuals who will be exposed to both HZE particles and low-LET radiation as they may be predisposed to increased late normal tissue damage.

Use of subjective and nonsubjective methodologies to evaluate lens radiation damage in exposed populations--an overview.

The general epidemiological acceptability of prevalence, or incidence, for assessing risk of radiation cataract development has dictated an almost exclusive dependence on cataract onset as a measure of cataractogenicity for given doses of radiation. The advent of instrumentation capable of acquiring images amenable to quantitative analyses offers the possibility of exploiting "relative opacification" as an added, if not exclusive, parameter. This development is particularly important in efforts to assess populations such as that in the Altai, which are temporally far removed from their exposure and among whom there exists a large subset with extant cataracts. The new technologies, Scheimpflug and retroillumination imaging, combined with the application of the appropriate analytical algorithms can not only provide quantitative and nonsubjective assessment of lens transparency, but also serve as a means to immortalize the state of the pathology at the time of acquisition. Highly relevant to the assessment of an aging exposed population is the use of lens epithelial fragments as potential dosimeters. The material is routinely available as a result of cataract extraction procedures and is amenable to the application of a modified micronucleus (MN) assay. The MN assay in the lens has tremendous advantages over its use in other tissues for a number of reasons, not least of which is that lens MNs are extremely long-lived. Given the relative ease of application and its potential as a radiation bioindicator, the lens MN assay should be considered in any follow-up of populations exposed to ionizing radiation.

Quantitative assessment of the cataractogenic potential of very low doses of neutrons.

We report on the prevalence and relative biological effectiveness (RBE) for various stages of lens opacification in rats induced by very low doses (2 to 250 mGy) of medium-energy (440 keV) neutrons, compared to those for X rays. Neutron doses were delivered either in a single fraction or in four separate fractions and the irradiated animals were followed for over 100 weeks. At the highest observed dose (250 mGy) and at early observation times, there was evidence of an inverse dose-rate effect; i.e., a fractionated exposure was more potent than a single exposure. Neutron RBEs relative to X rays were estimated using a non-parametric technique. The results were only weakly dependent on time postirradiation. At 30 weeks, for example, 80% confidence intervals for the RBE of acutely delivered neutrons relative to X rays were 8-16 at 250 mGy, 10-20 at 50 mGy, 50-100 at 10 mGy and 250-500 at 2 mGy. The results are consistent with the estimated neutron RBEs in Japanese A-bomb survivors, though broad confidence bounds are present in the Japanese results. Our findings are also consistent with data reported earlier for cataractogenesis induced by heavy ions in rats, mice, and rabbits. We conclude from these results that, at very low doses (<10 mGy), the RBE for neutron-induced cataractogenesis is considerably larger than the RBE of 20 commonly used, and use of a significantly larger value for calculating equivalent dose would be prudent.

Quantitative comparisons of continuous and pulsed low dose rate regimens in a model late-effect system.

PURPOSE: There is increasing interest and usage of pulsed low dose rate (PDR) brachytherapy, in which a single source is shuttled through the catheters of an implant, typically for about 10 min each hour. This study was designed to compare the late effects produced in various PDR regimens with those from the corresponding continuous low dose rate (CLDR) regimens. METHODS AND MATERIALS: A model late-responding system was used, namely, cataract induction in the rat lens. This system has the advantage of being highly quantifiable. The rats eyes were exposed to a total dose of 15 Gy either continuously over 24 h, or with three different PDR regimens, all with the same total dose and overall time. We addressed three questions: (a) are late effects increased when a CLDR regimen is replaced with 10-min pulses repeated every hour? (b) Are late effects increased if hourly 10-min pulses are replaced with 10-min pulses repeated every 4 h? (c) Are late effects increased if 10-min pulses are replaced with 100-s pulses? RESULTS: We found that the four regimens under test, continuous, 10-min pulses each hour, 10-min pulses every 4 h, and 100-s pulses every hour, showed no significant differences in cataractogenic potential, as estimated with the Wilcoxon-Gehan test. Power tests indicated that the experimental design was adequate to detect relatively small differences in cataractogenicity between regimens. CONCLUSIONS: The equality of late effects from CLDR and PDR in these experiments must imply that sublethal damage repair is quite slow in this model late-responding system, in agreement with trends observed in the clinic for sublethal damage repair of late sequelae. Such trends would suggest that PDR is unlikely to produce significantly worse late effects than the corresponding CLDR regimen, which is in agreement with early clinical data using PDR. Caution, however, is strongly recommended.

Use of non-subjective analysis of lens transparency in experimental radiation cataract research.

Historically the major impediment to radiation cataract follow-up has been the necessarily subjective nature of assessing the degree of lens transparency. This has spurred the development of instruments which produce video images amenable to digital analysis. One such system, the Zeiss Scheimpflug slit lamp measuring system (SLC), was incorporated into our ongoing studies of radiation cataractogenesis. It was found that the Zeiss SLC measuring system has high resolution and permits the acquisition of reproducible images of the anterior segment of the eye. Our results, based on about 650 images of lenses followed over a period of 91 weeks of radiation cataract development, showed that the changes in the light scatter of the lens correlated well with conventional assessment of radiation cataracts with the added advantages of objectivity, permanent and transportable records and linearity as cataracts become more severe. This continuous data acquisition, commencing with cataract onset, can proceed through more advanced stages. The SLC exhibits much greater sensitivity reflected in a continuously progressive severity thereby avoiding the artifactual plateaus in staging which occur using conventional scoring methods.

Radiation effects on late cytopathological parameters in the murine lens relative to particle fluence.

Lenses of mice irradiated with 250 MeV protons, 670 MeV/amu 20Ne, 600 MeV/amu 56Fe, 600 MeV/amu 93Nb and 593 MeV/amu 139La ions were evaluated by analyzing cytopathological indicators which have been implicated in the cataractogenic process. The LETs ranged from 0.40 keV/micrometer to 953 keV/micrometer and fluences from 1.31 10(3)/mm2 to 4.99 x 10(7)/mm2. 60Co gamma-rays were used as the reference radiation. The doses ranged from 10 to 40 cGy. The lenses were assessed 64 weeks post irradiation in order to observe the late effects of LET and dose on the target cell population of the lens epithelium. Our study shows that growth dependent pathological changes occur at the cellular level as a function of dose and LET. The shapes of the RBE-LET and RBE-dose curves are consistent with previous work on eye and other biological systems done in both our laboratory and others. The RBEmax's were estimated, for the most radiation cataract related cytological changes, MN frequency and MR disorganization, by calculating the ratio of the initial slopes of dose effect curve for various heavy ions to that of 60Co gamma-ray. For each ion studied, the RBEmax derived from micronucleus (MN) frequency is similar to that derived from meridional row (MR) disorganization, suggesting that heavy ions are equally efficient at producing each type of damage. Furthermore, on a per particle basis (particle/cell nucleus), both MN frequency and MR disorganization are LET dependent indicating that these classic precataractogenic indicators are multi-gene effects. Poisson probability analysis of the particle number traversing cell nuclei (average area = 24 micrometers2) suggested that single nuclear traversals determine these changes. By virtue of their precataractogenic nature the data on these endpoints intimate that radiation cataract may also be the consequence of single hits. In any case, these observations are consistent with the current theory of the mechanism of radiation cataractogenesis, which proposes that genomic damage to the epithelial cells surviving the exposure is responsible for opacification.

Modulating radiation cataractogenesis by hormonally manipulating lenticular growth kinetics.

There is considerable evidence that the lens epithelium is the primary site of injury leading to the development of cataracts following radiation exposure. That the damaged cells of the epithelium are the progenitors of the aberrantly differentiating fibers associated with the cataract is indisputable. So too is the observation that post-radiation proliferative activity in the lens epithelium is required for cataracts to develop. The natural hormonal regulation of lens epithelial mitotic activity in the frog offers the opportunity to alter the cell cycle of the lens epithelium in vivo, thus enabling the direct examination of the role of lenticular mitosis in the cytopathomechanism of radiation-induced cataracts. The cell cycle of the lens epithelium of northern leopard frogs was manipulated by hypophysectomy (to halt mitotic activity) and pituitary hormone administration (to stimulate baseline mitosis and reverse hypophysectomy-induced mitotic suppression). Animals were hypophysectomized, irradiated and injected with pituitary hormone replacement. Irradiated animals, irradiated animals + hormone replacement and irradiated hypophysectomized animals served as controls. Cataract development was evaluated by slit-lamp biomicroscopy and correlated with histologic determinations of mitotic index and meridional row disorganization on lens epithelial whole mounts. In another study, hypophysectomized-irradiated animals received varying concentrations of replacement hormone in an attempt to quantitatively modulate lens epithelial mitotic activity and determine the effect on cataractogenesis. It was found that irradiated-hypophysectomized (mitosis halted) frogs failed to develop opacities, while those with hormonal replacement (mitosis reinstated) developed cataracts.(ABSTRACT TRUNCATED AT 250 WORDS)

Neuritogenesis: a model for space radiation effects on the central nervous system.

Pivotal to the astronauts' functional integrity and survival during long space flights are the strategies to deal with space radiations. The majority of the cellular studies in this area emphasize simple endpoints such as growth related events which, although useful to understand the nature of primary cell injury, have poor predictive value for extrapolation to more complex tissues such as the central nervous system (CNS). In order to assess the radiation damage on neural cell populations, we developed an in vitro model in which neuronal differentiation, neurite extension, and synaptogenesis occur under controlled conditions. The model exploits chick embryo neural explants to study the effects of radiations on neuritogenesis. In addition, neurobiological problems associated with long-term space flights are discussed.

The influence of dose, dose-rate and particle fragmentation on cataract induction by energetic iron ions.

Because activities in space necessarily involve chronic exposure to a heterogeneous charged particle radiation field it is important to assess the influence of dose-rate and the possible modulating role of heavy particle fragmentation on biological systems. Using the well-studied cataract model, mice were exposed to plateau 600 MeV/amu 56Fe ions either as acute or fractionated exposures at total doses of 5 - 504 cGy. Additional groups of mice received 20, 360 and 504 cGy behind 50 mm of polyethylene, which simulates body shielding. The reference radiation consisted of 60Co gamma radiation. The animals were examined by slit lamp biomicroscopy over their three year life spans. In accordance with our previous observations with heavy particles, the cataractogenic potential of the 600 MeV/amu 56Fe ions was greater than for low-LET radiation and increased with decreasing dose relative to gamma-rays. Fractionation of a given dose of 56Fe ions did not reduce the cataractogenicity of the radiation compared to the acute regimen. Fragmentation of the beam in the polyethylene did not alter the cataractotoxicity of the ions, either when administered singly or in fractions.

Non-subjective cataract analysis and its application in space radiation risk assessment.

Experimental animal studies and human observations suggest that the question is not whether or not prolonged space missions will cause cataracts to appear prematurely in the astronauts, but when and to what degree. Historically the major impediment to radiation cataract follow-up has been the necessarily subjective nature of assessing the degree of lens transparency. This has spurred the development of instruments which produce video images amenable to digital analysis. One such system, the Zeiss Scheimpflug slit lamp measuring system (SLC), was incorporated into our ongoing studies of radiation cataractogenesis. It was found that the Zeiss SLC measuring system has high resolution and permits the acquisition of reproducible images of the anterior segment of the eye. Our results, based on about 650 images of the rats lens, and followed over a period of 91 weeks of radiation cataract development, showed that the integrated optical density (IOD) of the lens correlated well with conventional assessment with the added advantages of objectivity, permanent and transportable records and linearity as cataracts become more severe. This continuous data acquisition, commencing with cataract onset, can proceed through more advanced stages. The SLC exhibits much greater sensitivity reflected in a continuously progressive severity despite the artifactual plateaus in staging which occur using conventional scoring methods. Systems such as the Zeiss SLC should be used to monitor astronauts frequent visits to low earth orbit to obtain a longitudinal data-base on the influence of this activity on the lens.

Accelerated heavy ions and the lens. IX. Late effects of LET and dose on cellular parameters in the murine lens.

Lenses of mice irradiated with 250 MeV protons, 670 MeV/amu 20Ne, 600 MeV/amu 56Fe, 350 MeV/amu 56Fe, 600 MeV/amu 93Nb or 593 MeV/amu 139La ions were evaluated by analysing cytopathological indicators which have been implicated in the cataractogenic process. The LETs ranged from 0.39 to 953 keV/microns and the fluences from 1.31 x 10(3)/mm2 to 5.12 x 10(7)/mm2. The lenses were assessed 64 weeks post-irradiation in order to observe the late effects of LET and dose on the target cell population of the lens' epithelium. Our studies showed that growth-dependent pathological changes occurred at the cellular level as a function of dose and LET. For a given particle dose, as the LET rose, the number of abnormal mitotic figures, micronuclei frequency, and the disorganization of meridional rows increased to a maximum and then reached a plateau or decreased. For particles of the same LET, the severity of meridional rows disorganization and micronuclei frequency increased with increasing dose. The numbers of cells surviving at late times post-irradiation were comparable with those of controls. In addition, the cellular density was similarly unaffected. These observations are consistent with the current theory of the mechanism of radiation cataractogenesis which posits that genomic damage to the epithelial cells surviving the exposure is responsible for opacification.

Accelerated heavy particles and the lens. VIII. Comparisons between the effects of acute low doses of iron ions (190 keV/microns) and argon ions (88 keV/microns).

The nature of the RBE-LET relationship for radiation-induced effects in vivo is not well known in the high-LET range above about 100 keV/microns. Here, we compare the cataractogenic effects of acute doses of 190 keV/microns iron ions on the eyes of rats with those of 88 keV/microns argon ions. The RBEs of the two radiations cannot be distinguished statistically, both being between 50 and 200 at our lowest dose of 0.01 Gy and decreasing to between 2 and 14 at our highest dose of 0.5 Gy; these values are consistent with results obtained in vivo, both for cataractogenesis and for oncogenesis. For this end point, therefore, the RBE-LET relationship probably varies very slowly between 88 and 190 keV/microns. On the basis of these studies with acute doses of 88 and 190 keV/microns particles, the detailed distribution in LET of the very high-LET galactic cosmic-ray dose to which astronauts in deep space are exposed may not be critical for the prediction of biological hazard. Such a conclusion might simplify the task of high-LET radiation risk estimation in space.

Accelerated heavy particles and the lens. VII: The cataractogenic potential of 450 MeV/amu iron ions.

PURPOSE: To determine the cataractogenic potential dose of high velocity iron ions as a fixation of dose administered singly or fractionated. The dose is critical to risk assessment and to theories of radiation action and cataractogenesis. METHODS: Twenty-eight-day-old rats were examined by slit-lamp biomicroscopy on a weekly-bi-weekly basis for more than 2 yr after radiation exposure. For the acute exposure study doses of 1, 2, 5, 25, and 50 cGy were evaluated. The fractionated regimens involved total doses of 2, 25, and 50 cGy. The reference radiation consisted of 50, 100, 200, or 700 cGy of 250 kilovolt (peak) x-rays. RESULTS: In accordance with previous findings in the rat using 570 MeV/amu 40Ar ions, the relative biologic effectiveness increased rapidly with decreasing dose, reaching values as high as 100. Unlike 40Ar ions, fractionation of the 56Fe doses did not produce a consistent enhancement at any of the doses examined. CONCLUSIONS: The data support the previous findings of a high cataractogenic potential for high linear energy transfer (LET) radiation. The effectiveness for the production of cataracts increases with decreasing dose relative to x-rays and is independent of dose protraction. Although the present study did not reveal a consistent enhancement of effect when the ions were applied in fractions, the results are consistent with at least one theory of the inverse dose-rate effect observed for high-LET radiation.

Do heavy ions cause microlesions in cell membranes?

Heavy ions are a hazard in manned deep space missions. It has been theoretically postulated that when they interact with cells, localized damage in the forms of "microlesions" may occur. Purported morphological evidence of these lesions, however, has not been confirmed in the most extensively studied tissue so far, the cornea. Recent morphological evidence from rat corneas demonstrated that holes in membranes do not form as consequence of heavy ion irradiation. This does not mean, however, that some other form of damage is excluded. For example such damage may be physiological in nature, impairing the ability of cells or tissues to function properly. In order to uncover any physiological effects, we investigated the microlesion question by monitoring the electrical potential difference across the endothelium of rat corneas in vitro before, during, and after irradiation. When the corneas were exposed to 1 Gy of 56Fe ions (450 and 600 MeV/a.m.u.), we detected no effect on this parameter. These results suggest that direct physical damage to cell membranes, as predicted by the microlesion theory, does not take place.

Evidence of genotoxic damage in human cataractous lenses.

Lens epithelial fragments (tags) recovered from individuals during routine cataract extraction have been assessed for cellular changes reflective of genotoxic damage. A high percentage of tags exhibited a population of micronucleated and polyploid cells. The presence and number of micronuclei (MN) in the epithelia of cataract patients appears to be independent of age and sex. However, a large number of MN in the epithelial cells of some individuals strongly suggests a history of compromised genomic integrity. While the study was not designed to define the role of DNA damage in the development of cataracts or to monitor human populations at risk of exposure to exogenous mutagens/cataractogens, the potential of the methodology to address each is demonstrated.

Accelerated heavy particles and the lens. VI. RBE studies at low doses.

We report on the prevalence, hazard, and relative biological effectiveness (RBE) for various stages of lens opacification in rats induced by very low doses of fast argon ions of LET 88 keV/microns, compared to those for X rays. Doses of argon ions from 0.01 to 0.25 Gy were used and RBEs of these ions relative to X rays estimated using a nonparametric technique. At the end of the follow-up period, which encompasses a significant fraction of the animals' lifetime, 90% confidence intervals for the RBE of the argon ions relative to X rays were 4-8 at 0.25 Gy, 10-40 at 0.05 Gy, and 50-100 at 0.01 Gy. Our results are consistent with the point-estimate neutron RBEs in Japanese A-bomb survivors, though broad confidence bounds are present in the Japanese results. If a reasonable extrapolation to higher doses is used, our results are also consistent with data reported earlier at higher doses for argon-ion cataractogenesis in rats, mice, and rabbits. We conclude from these results that at very low doses the RBE for cataractogenesis from HZE particles in space is considerably more than 20, and use of a quality factor of at least 50 would be prudent.

Neutron effects on the lens.

Few in vivo systems have received the investigative attention or have provided the grist for our understanding of basic radiobiological principles as have the lens and the cataract. From Roentgen's time the lens has been recognized as a "biological dosimeter" for gauging radiation response. Its advantages range from its in vivo status to its qualification as an integrated tissue. From the time of the Hiroshima/Nagasaki experience, there has been some urgency in attempting to understand the breadth of neutron-radiation effects on humans. The major obstacle has been our understanding of the doses which were received by the individuals who express the damage. The majority of the work has been derived from experimental animals and findings related to photons: X and gamma rays. Cataractogenesis provides insights in terms of not only ocular radiopathy but also the basic mechanisms of the action of radiation. Often referred to as the "classic" nonstochastic radiation effect, it is becoming increasingly clear that the suggestion of a threshold reflects the limitations imposed on expression by the life span. Thus the primary damage which appears to be a somatic mutation is fully stochastic. This being the case, it is not surprising that, as is the case for simpler systems, the RBEs for cataracts following neutron exposure are significantly higher than for X rays, and that there is evidence for an inverse dose-rate effect in their production. This presentation focuses on these data and on the merits of the lens for the assessment of neutron effects on humans, the existing data for known dose levels in the human population, and the confounding issues associated with extrapolation from experimental work. Data from Western sources as well as those from the USSR are presented.

Rapid deterioration of lens fibers in GSH-depleted mouse pups.

Lens opacities developed within 48-72 hr in mice that received a series of eight injections of L-buthionine sulfoximine, a specific inhibitor of glutathione (GSH) biosynthesis, on postnatal days 8 and 9. Initial histopathologic features consisted of swollen fibers in the central anterior cortex and displacement of cell nuclei from the bow region to the posterior cortex. These aberrations suggest early fiber cell membrane and/or cytoskeletal dysfunction. A massive wave of fiber cell lysis then engulfed the entire lens cortex and nucleus within 24 hr and left only epithelial cells intact, suggesting a concerted mechanism of cataract generation. The acellular core of the mature cataract seen on postnatal day 16 consisted of a granular matrix in which pycnotic and fragmented cell nuclei were located near the terminus of the lens epithelium. The epithelium displayed increased mitotic activity and meridional row disorganization. During the next two weeks, rapid regeneration of lens fibers, displacement of the acellular necrotic cytoplasm to the center and rear of the lens, and vacuole formation were observed. As new fibers were differentiated, partial regeneration of the bow was seen. However, the cataract was irreversible.